Atomic force microscopy (AFM) has become an important tool in nanoscale science and applications. A micro-sized passive cantilever structure in a general AFM arrangement is used to measure interaction forces between a sharp needle integrated with the cantilever and a surface that the needle interacts with. The cantilever is moved over the surface by the help of a piezo actuator. In the meantime, the force acting on the cantilever causes bending on the cantilever depending on its spring constant. Bending is measured by optical means. FIG. 1 conceptually shows an AFM cantilever scanning the surface, the closed-loop control system that controls operation thereof and detection optics.
AFM is the most commonly used technology for single-molecule mechanics measurements thanks to its versatility. Commercially available AFM cantilevers provide force resolution of few picoNewtons (pN) in liquid. Cantilevers should be actuated within a specific speed range for AFM-based dynamic force spectroscopy applications. Higher dynamic range for actuation and detection is required along with higher force resolution for improved dynamic force spectroscopy. The pulling speed of the cantilever by using a piezo actuator determines the molecular loading rate in a conventional AFM system.
Hydrodynamic drag force on the moving cantilever increases with the pulling speed. Drag force may reach the level of biomolecular forces, which is desired to be measured, even at moderate speed levels, i.e., a few tens of μm/s for most of the cantilevers in use. However higher pulling speed capability is required for complete understanding of energy landscapes. In the light of these, the need for novel force sensors and technologies that allow high loading rates with minimal hydrodynamic drag forces on the force sensors can be put in a proper context.
Magnetic micro-beads are frequently used for actuating biomolecules by magnetic particles. The present invention, which enables manipulation of biomolecules for investigating their conformational behavior with high precision along multiple degrees of freedom, is fundamentally different from the approaches used in conventional magnetic manipulation systems in which magnetic beads are used. The magnetic micro-beads are typically made of iron oxide particles covered by polymer matrices. A spherical single domain magnetite particle with a diameter of 100 nm cannot exert forces in excess of tens of picoNewtons given practical limitations on the generation of magnetic field gradients, and is incapable of generating torque due to its geometry. This is a significant limitation in their use. The size of the particles may be increased to increase the attainable force levels; however, surface area of magnetic particles with diameters larger than 1 μm is much larger than an area to which a single biomolecule can bind. Therefore, their use in single-molecule measurements is limited. High aspect ratio magnetic nanowires address this bottleneck. Magnetic nanowires exhibit larger magnetic moments as compared to the beads because of their higher volume and their inherent magnetic shape anisotropy. Upon comparing the attainable force levels, it was observed that nickel magnetic nanowires outperformed magnetic beads of the same volume by a factor of two. Moreover, when increasing the volume of nanowires, the size of its tip binding to the molecules does not have to be increased. Volume of a nanowire can be increased by extending its length while keeping its diameter small enough so that it can still interact with single molecules. Additionally, nanowires can also generate torque thanks to their geometric forms.
The International patent application document no. WO2011029592, an application in the state of the art, discloses a magnetic manipulation device for magnetic elements.
The structure disclosed in U.S. Pat. document No. U.S. Pat. No. 8,479,309B2, an application in the state of the art, is a novel AFM cantilever. An improved measurement cantilever is produced by means of a nano-needle integrated to an AFM cantilever and it is disclosed that by means of this cantilever improvements are achieved in the measurements conducted in liquid. This method is aimed for developing a new cantilever, and the nano-needle arranged on the cantilever only provides a new geometry to the cantilever. The innovation disclosed by the biomolecular measurement system of the present invention is combination of magnetic nanowires with biomolecules and using the nanowires as actuators. In the bimolecular measurement system of the present invention, the nanowires are integral part of the measurement system and they provide magnetic actuation capability.
The document titled “New fabrication methods and measurement techniques enable development of nanoscale bimorph actuators” (E. H. Yang, 23 Jun. 2010, SPIE Newsroom. DOI: 10.1117/2 1201006.002602) presents nano-scale actuators produced with bimorph structures. The feature of bimorph structures is that they bend under temperature difference. By means of this feature, it is possible to exert force at micro and nano-scale. The nano-structures are used to exert force on biological structures. The nanowires used in the biomolecular measurement system of the present invention are the actuators of the AFM system. The nanowires described in the present invention are magnetic actuators and differ from the nano-structures of bimorph actuators in terms of both function and structure.
The article titled “Control of tip-to-sample distance in atomic force microscopy: a dual-actuator tip-motion control scheme” (Jeong Y, Jayanth G R, Menq C H., Rev Sci Instrum. 2007 September; 78(9):093706) describes actuation of a standard AFM cantilever by a magnetic particle adhered on the AFM cantilever. The mechanical structure moved in this system is the cantilever itself and the magnetic particle is fixed to the cantilever. In the biomolecular measurement system of the present invention, the moving structure is nanowire whereas the cantilever is fixed. The cantilever moves only as a result of biomolecular interaction forces. It does not move due to the signal applied to the actuator. Thus, low noise and high stability measurements can be performed.
The document titled “Electromagnetically Actuated Cantilevers Using Magnetic Micropillars for Atomic Force Microscopy” (A Fakhraee, N Shamsudhin, S Sevim, A Lindo, S Pane, O Ergeneman, H Torun, B Nelson, IEEE International Magnetics Conference, May 4-8 2014, Dresden, Germany) discloses that the cantilever is moved by the magnetic forces in an application similar to the structure described by the document titled “Control of tip-to-sample distance in atomic force microscopy: a dual-actuator tip-motion control scheme”. A micropillar is adhered onto the cantilever. The actuation method formed by an electromagnet is different from the method used in the biomolecular measurement system of the present invention. In the present invention, while the cantilever remains fixed, the magnetic nanowires are actuated by the actuator signal. Furthermore, the nanowires in the present invention are much smaller than the micropillars described in the mentioned document.